While there is a wide variety of methods used for the electrotransformation of Lactobacillus spp., they all share a typical format.

While there is a wide variety of methods used for the electrotransformation of Lactobacillus spp., they all share a typical format.

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*Dilution: This is the amount of overnight culture to add to new gorwth media

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*Dilution: This determines the amount of overnight culture to add to new growth media

*Growth: These are the conditions used to grow the cells on the day you plan to transform.

*Growth: These are the conditions used to grow the cells on the day you plan to transform.

*Washing: Cells are repeatedly centrifuged and resuspended in a solution of some type.

*Washing: Cells are repeatedly centrifuged and resuspended in a solution of some type.

*Buffer: This is what is used for the final resuspension and electroporation.

*Buffer: This is what is used for the final resuspension and electroporation.

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*Concentration: This determines how much Electroporation buffer to add relative to the volume of the growth media.

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*Voltage: How hard to shock em.

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*Recovery: What to do with your cells after the shock.

This page will list many of these protocols in a standard format so that you can compare them and choose the one that works for you. In some cases an apples to apples comparison may not be acceptable (e.g. the differences in electroporators can be dramatic) so be sure to check the actual paper.

This page will list many of these protocols in a standard format so that you can compare them and choose the one that works for you. In some cases an apples to apples comparison may not be acceptable (e.g. the differences in electroporators can be dramatic) so be sure to check the actual paper.

# Centrifugation at 4000 rpm for 2 minutes was sufficient to pellet the competent cells to give a clear supernatant.

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#Centrifugation at 4000 rpm for 2 minutes was sufficient to pellet the competent cells to give a clear supernatant.

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# Most of these papers use the Bio-Rad Gene Pulser and have time constants in the range of 9-10 ms with the lower voltages (7-9kV/cm). If you're using a preset electroporator (like the Eppendorf 2510) you should try a higher voltage (~1200) as your time constand will be in thee range of 5-6 ms. If you're using PEG in your electroporation buffer expect your time constant to be significantly lower.

-

#Most of these papers use the Bio-Rad Gene Pulser and have time constants in the range of 9-10 ms with the lower voltages (7-9kV/cm). If you're using a preset electroporator (like the Eppendorf 2510) you should try a higher voltage (~1200) as your time constand will be in thee range of 5-6 ms. If you're using PEG in your electroporation buffer expect your time constant to be significantly lower.

+

# If you get ZERO TRANSFORMANTS after trying multiple methods, chances are that your plasmid is incompatible with the strain you're transforming. This page exists because this happened to me; and after I had tried ALL the protocols listed here, I got a new plasmid and it worked beautifully -- Mike.

==References==

==References==

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This protocol is based mostly on the protocols described by Thompson and Collins (1996) and Mason et al. (2004) but other literature are also listed.

Thompson, K. and M. A. Collins (1996). "Improvement in electroporation efficiency for Lactobacillus plantarum by the inclusion of high concentrations of glycine in the growth medium." Journal of Microbiological Methods 26(1-2): 73-79.

*Thompson, K. and M. A. Collins (1996). "Improvement in electroporation efficiency for ''Lactobacillus plantarum'' by the inclusion of high concentrations of glycine in the growth medium." ''Journal of Microbiological Methods'' 26(1-2): 73-79.

Overview

While there is a wide variety of methods used for the electrotransformation of Lactobacillus spp., they all share a typical format.

Dilution: This determines the amount of overnight culture to add to new growth media

Growth: These are the conditions used to grow the cells on the day you plan to transform.

Washing: Cells are repeatedly centrifuged and resuspended in a solution of some type.

Buffer: This is what is used for the final resuspension and electroporation.

Concentration: This determines how much Electroporation buffer to add relative to the volume of the growth media.

Voltage: How hard to shock em.

Recovery: What to do with your cells after the shock.

This page will list many of these protocols in a standard format so that you can compare them and choose the one that works for you. In some cases an apples to apples comparison may not be acceptable (e.g. the differences in electroporators can be dramatic) so be sure to check the actual paper.

Notes

Centrifugation at 4000 rpm for 2 minutes was sufficient to pellet the competent cells to give a clear supernatant.

Most of these papers use the Bio-Rad Gene Pulser and have time constants in the range of 9-10 ms with the lower voltages (7-9kV/cm). If you're using a preset electroporator (like the Eppendorf 2510) you should try a higher voltage (~1200) as your time constand will be in thee range of 5-6 ms. If you're using PEG in your electroporation buffer expect your time constant to be significantly lower.

If you get ZERO TRANSFORMANTS after trying multiple methods, chances are that your plasmid is incompatible with the strain you're transforming. This page exists because this happened to me; and after I had tried ALL the protocols listed here, I got a new plasmid and it worked beautifully -- Mike.

Thompson, K. and M. A. Collins (1996). "Improvement in electroporation efficiency for Lactobacillus plantarum by the inclusion of high concentrations of glycine in the growth medium." Journal of Microbiological Methods 26(1-2): 73-79.